Back to EveryPatent.com
| United States Patent |
5,258,126
|
|
Pall
, et al.
|
*
November 2, 1993
|
Method for obtaining platelets
Abstract
A method is provided for processing donated blood, particularly a
platelet-containing solution such as platelet-containing plasma, involving
separating blood into a red cell containing sediment layer and a
supernatant layer, and passing the supernatant layer through a filter
until the filter is blocked, thereby leaving platelets to be harvested.
The preferred filter comprises a porous medium having a plurality of zones
of progressively increasing density.
| Inventors:
|
Pall; David B. (Roslyn Estates, NY);
Gsell; Thomas C. (Glen Cove, NY)
|
| Assignee:
|
Pall Corporation (Glen Cove, NY)
|
| [*] Notice: |
The portion of the term of this patent subsequent to October 6, 2009
has been disclaimed. |
| Appl. No.:
|
846587 |
| Filed:
|
March 5, 1992 |
| Current U.S. Class: |
210/767; 210/782; 210/787; 210/789; 210/806 |
| Intern'l Class: |
B01D 037/00; B01D 021/26 |
| Field of Search: |
210/767,782,787,789,806
422/41,44
604/4,5,6,406
|
References Cited
U.S. Patent Documents
| 2834730 | May., 1958 | Painter, Jr. et al. | 210/504.
|
| 2956899 | Oct., 1960 | Cline | 522/144.
|
| 3111424 | Nov., 1963 | LeClair | 522/144.
|
| 3268622 | Aug., 1966 | Stanton et al. | 522/144.
|
| 3274294 | Sep., 1966 | Stanton et al. | 522/144.
|
| 3448041 | Jun., 1969 | Swank.
| |
| 3565780 | Feb., 1971 | Zimmrman | 522/144.
|
| 3593854 | Jul., 1971 | Swank | 210/436.
|
| 3663374 | May., 1972 | Moyer et al.
| |
| 3765536 | Oct., 1973 | Rosenberg | 210/446.
|
| 3877978 | Apr., 1975 | Kremen et al. | 210/508.
|
| 3905905 | Sep., 1975 | O'Leary et al. | 210/436.
|
| 3935110 | Jan., 1976 | Schmid et al. | 210/456.
|
| 3935111 | Jan., 1976 | Bentley | 210/446.
|
| 3954621 | May., 1976 | Etani et al. | 210/416.
|
| 4009714 | Mar., 1977 | Hammer | 210/436.
|
| 4009715 | Mar., 1977 | Forberg et al. | 210/455.
|
| 4073732 | Feb., 1978 | Lauer et al. | 210/491.
|
| 4081402 | Mar., 1978 | Levy et al. | 424/78.
|
| 4087363 | May., 1978 | Rosemeyer et al. | 210/448.
|
| 4092246 | May., 1978 | Kummer | 210/504.
|
| 4115277 | Sep., 1978 | Swank | 210/436.
|
| 4116845 | Sep., 1978 | Swank | 210/446.
|
| 4157967 | Jun., 1979 | Meyst et al. | 210/489.
|
| 4170056 | Oct., 1979 | Meyst et al. | 210/446.
|
| 4229309 | Oct., 1980 | Cheng et al. | 428/489.
|
| 4246107 | Jan., 1981 | Takenaka et al. | 210/259.
|
| 4283289 | Aug., 1981 | Meyst et al. | 210/448.
|
| 4294594 | Oct., 1981 | Sloane, Jr. et al. | 210/446.
|
| 4330410 | May., 1982 | Takenaka et al. | 210/767.
|
| 4334901 | Jun., 1982 | Ayes et al. | 55/487.
|
| 4370381 | Jan., 1983 | Horikoshi et al. | 210/508.
|
| 4376675 | Mar., 1983 | Perrotta | 210/509.
|
| 4416777 | Nov., 1983 | Kuroda et al. | 210/446.
|
| 4422939 | Dec., 1983 | Sharp et al. | 210/445.
|
| 4443492 | Apr., 1984 | Roller | 604/378.
|
| 4459210 | Jul., 1984 | Murakami et al. | 428/398.
|
| 4476023 | Oct., 1984 | Horikoshi et al. | 210/446.
|
| 4477575 | Oct., 1984 | Vogel et al. | 210/509.
|
| 4534757 | Aug., 1985 | Geller | 604/85.
|
| 4596657 | Jun., 1986 | Wisdom | 210/206.
|
| 4608173 | Aug., 1986 | Watanabe et al. | 210/502.
|
| 4617124 | Oct., 1986 | Pall et al. | 210/508.
|
| 4618533 | Oct., 1986 | Steuck | 428/315.
|
| 4620932 | Nov., 1986 | Howery | 210/490.
|
| 4636312 | Jan., 1987 | Willis | 210/416.
|
| 4701267 | Oct., 1987 | Watanabe et al. | 210/491.
|
| 4702947 | Oct., 1987 | Pall et al. | 210/508.
|
| 4753739 | Jun., 1988 | Noble | 210/787.
|
| 4786603 | Nov., 1988 | Wielinger et al. | 210/500.
|
| 4816224 | Mar., 1989 | Vogel et al. | 210/767.
|
| 4833087 | May., 1989 | Hinckley | 422/101.
|
| 4855063 | Aug., 1989 | Carmen et al. | 210/749.
|
| 4880548 | Nov., 1989 | Pall et al. | 210/767.
|
| 4883764 | Nov., 1989 | Kloepfer | 422/56.
|
| 4909949 | Mar., 1990 | Harmony et al. | 210/787.
|
| 4923620 | May., 1990 | Pall | 210/767.
|
| 4925572 | May., 1990 | Pall | 210/767.
|
| 4933092 | Jun., 1990 | Aunet et al. | 210/510.
|
| 4936998 | Jun., 1990 | Nishimura et al. | 210/496.
|
| 4987085 | Jan., 1991 | Allen et al. | 422/56.
|
| 5100564 | Mar., 1992 | Pall et al. | 210/767.
|
| 5152905 | Oct., 1992 | Pall et al. | 210/767.
|
| Foreign Patent Documents |
| 114281 | Aug., 1984 | EP.
| |
| 370584 | May., 1990 | EP.
| |
| 446713 | Sep., 1991 | EP.
| |
| 455215 | Nov., 1991 | EP.
| |
| 2735179 | Feb., 1978 | DE.
| |
| 2511872 | Mar., 1983 | FR.
| |
| 9104088 | Apr., 1991 | WO.
| |
| 1501665 | Feb., 1978 | GB.
| |
| 2017713 | Oct., 1979 | GB.
| |
| 2231282 | Nov., 1990 | GB.
| |
Primary Examiner: Dawson; Robert A.
Assistant Examiner: Kim; Sun Uk
Attorney, Agent or Firm: Leydig, Voit & Mayer
Parent Case Text
This application is a continuation-in-part application of U.S. Ser. No.
07/609,574, filed Nov. 6, 1990, now issued U.S. Pat. No. 5,152,905 which
is a continuation-in-part of U.S. Ser. No. 07/405,977, filed Sep. 12, 1989
(now abandoned); and of U.S. Ser. No. 07/609,654, filed Nov. 6, 1990, now
issued U.S. Pat. No. 5,100,564.
Claims
What is claimed is:
1. A method of harvesting platelets from a platelet-containing solution
comprising:
separating the platelet-containing solution into a supernatant layer and a
sediment layer containing red cells; and
passing the supernatant layer through a filter until red cells block the
filter, said filter including a porous medium having zones of different
density.
2. The method of claim 1 wherein passing the supernatant layer through the
porous medium comprises passing platelet-rich plasma through the porous
medium.
3. The method of claim 1 wherein said filter comprises a porous medium
having a CWST greater than about 70 dynes/cm.
4. The method of claim 2 wherein passing the supernatant layer through a
filter further comprises passing the supernatant layer through a filter
having a flow area of about 3 cm.sup.2 to about 8 cm.sup.2.
5. The method of claim 1 wherein passing the supernatant layer through a
filter further comprises passing the supernatant layer through a filter
having a density range in an upstream portion from about 0.18 g/cc to
about 0.23 g/cc, and a density range in the downstream portion from about
0.23 g/cc to about 0.40 g/cc.
6. The method of claim 5 wherein passing the supernatant layer through a
porous medium having zones of different density comprises passing the
supernatant layer through at least two zones of different density.
7. The method of claim 6 wherein passing the supernatant layer through
zones of different density comprises passing the supernatant layer through
zones of successively higher density.
8. The method of claim 7 wherein passing the supernatant layer through at
least two zones of different density comprises passing the supernatant
layer through an upstream zone including a density range from about 0.1
g/cc to about 0.2 g/cc, through an intermediate zone including a density
range from about 0.20 g/cc to about 0.25 g/cc, and through a downstream
zone including a density range from about 0.23 g/cc to about 0.40 g/cc.
9. A method for treating blood comprising:
centrifuging blood to create a supernatant layer and a sediment layer; and
passing the supernatant layer of the centrifuged blood through a porous
medium until red blood cells block the porous medium, said filter
including a porous medium having zones of different density.
10. The method of claim 9 further comprising collecting the supernatant
layer passing through the porous medium.
Description
TECHNICAL FIELD OF THE INVENTION
The present invention relates to a method and apparatus for processing
blood donated for the purpose of therapeutic transfusion of blood
components and, particularly, to improved methods and apparatuses for
harvesting platelets from donated whole blood.
BACKGROUND OF THE INVENTION
The development of plastic blood collection bags facilitated the separation
of donated whole blood into its various components, e.g., platelet
concentrate (hereinafter "PC"), packed red cells (hereinafter "PRC"), and
plasma, thereby making platelet concentrates available as a transfusion
product. The separation of a single unit of donated whole blood, about 450
milliliter in USA practice, into its components is typically accomplished
by use of differential sedimentation.
A typical procedure used in the United States, the
citrate-phosphate-dextrose-adenine (CPDA-1) system, utilizes a series of
steps to separate donated blood into three components, each component
having substantial therapeutic and monetary value. The procedure typically
utilizes a blood collection bag which is integrally attached via tubing to
at least one, and preferably two or more, satellite bags. Whole blood may
be thus collected and processed as follows:
(1) The donated whole blood is collected from the donor's vein directly
into the blood collection bag which contains the nutrient and
anti-coagulant containing CPDA-1.
(2) The blood collection bag is centrifuged together with its satellite
bags, thereby concentrating the red cells as packed red cells (hereinafter
PRC) in the lower portion of the blood collection bag and leaving in the
upper portion of the bag a suspension of platelets in clear plasma, which
is known as platelet-rich plasma (PRP).
(3) The blood collection bag is transferred, with care not to disturb the
interface between the supernatant PRP layer and the sedimented PRC layer,
into a device known as a "plasma extractor" which comprises an opaque back
plate and a transparent front plate; the two plates are hinged together at
their lower ends and spring biased toward each other such that a pressure
of about 90 millimeters of mercury is developed within the bag.
With the blood collection bag positioned between the two plates, a valve or
seal in the tubing is opened allowing the supernatant PRP to flow into a
first satellite bag. As the PRP flows out of the blood collection bag, the
interface with the PRC rises. The operator closely observes the position
of the interface as it rises and clamps off the connecting tube when in
his judgment as much PRP has been transferred as is possible, consistent
with allowing no red cells to enter the first satellite bag. This is a
time consuming operation during which the operator must visually monitor
the bag and judiciously and arbitrarily ascertain when to shutoff the
connecting tube. The blood collection bag, now containing only PRC, may be
detached and stored at 4.degree. C. until required for transfusion into a
patient, or a valve or seal in the flexible tubing may be opened so that
the PRC may be transferred to a satellite bag using either the pressure
generated by the plasma extractor apparatus, or by placing the blood
collection apparatus in a pressure cuff, or by elevation to obtain gravity
flow.
(4) The PRP-containing satellite bag, together with another satellite bag,
is then removed from the extractor and centrifuged at an elevated G force
with the time and speed adjusted so as to concentrate the platelets into
the lower portion of the PRP bag. When centrifugation is complete, the PRP
bag contains sedimented platelets in its lower portion and clear plasma in
its upper portion.
(5) The PRP bag is then placed in the plasma extractor, and most of the
clear plasma is expressed into the other satellite bag, leaving the PRP
bag containing only sedimented platelets in about 50 ml of plasma; in a
subsequent step, this platelet composition is dispersed to make PC. The
PRP bag, now containing a PC product, is then detached and stored for up
to five days at 20.degree.-22.degree. C., until needed for a transfusion
of platelets. For use with adult patients, the platelets from 6-10 donors
are, when required, pooled into a single platelet transfusion.
(6) The plasma in the other satellite bag may itself be transfused into a
patient, or it may be separated by complex processes into a variety of
valuable products.
Commonly used systems other than CPDA-1 include Adsol, Nutricell, and
SAG-M. In these latter systems, the collection bag contains only
anticoagulant, and the nutrient solution may be preplaced in a satellite
bag. This nutrient solution is transferred into the PRC after the PRP has
been separated from the PRC, thereby achieving a higher yield of plasma
and longer storage life for the PRC.
With the passage of time and accumulation of research and clinical data,
transfusion practices have changed greatly. One aspect of current practice
is that whole blood is rarely administered; rather, patients needing red
blood cells are given packed red cells, patients needing platelets are
given platelet concentrate, and patients needing plasma are given plasma.
For this reason, the separation of blood into components has substantial
therapeutic and monetary value. This is nowhere more evident than in
treating the increased damage to a patient's immune system caused by the
higher doses and stronger drugs now used during chemotherapy for cancer
patients. These more aggressive chemotherapy protocols are directly
implicated in the reduction of the platelet content of the blood to
abnormally low levels; associated internal and external bleeding
additionally requires more frequent transfusions of PC, and this has
caused platelets to be in short supply and has put pressure on blood banks
to increase platelet yield per unit of blood.
Blood bank personnel have responded to the increased need for blood
components by attempting to increase PC yield in a variety of ways,
including attempting to express more PRP prior to stopping flow from the
blood collection bag. This has often proved to be counterproductive in
that the PRP, and the PC subsequently extracted from it, are frequently
contaminated by red cells, giving a pink or red color to the normally
light yellow PC. The presence of red cells in PC is so highly undesirable
that pink or red PC is frequently discarded, or subjected to
recentrifugation, both of which increase operating costs.
The method and apparatus of the present invention alleviate the
above-described problems and, in addition, provide a higher yield of
superior quality PC.
In addition to the three above-listed components, whole blood contains
white blood cells (known collectively as leucocytes) of various types, of
which the most important are granulocytes and lymphocytes. White blood
cells provide protection against bacterial and viral infection.
The transfusion of blood components which have not been leucocyte-depleted
is not without risk to the patient receiving the transfusion. Chills,
fever, and allergic reactions may occur in patients receiving acute as
well as chronic platelet therapy. Repeated platelet transfusions
frequently lead to alloimmunization against HLA antigens, as well as
platelet specific antigens. This, in turn, decreases responsiveness to
platelet transfusion. Leucocytes contaminating platelet concentrates,
including granulocytes and lymphocytes, are associated with both febrile
reactions and alloimmunization, leading to platelet transfusion
refractoriness. Another life-threatening phenomenon affecting heavily
immunosuppressed patients is Graft Versus Host Disease. In this clinical
syndrome, donor lymphocytes transfused with the platelet preparations can
launch an immunological reaction against the transfusion recipient with
pathological consequences. Some of these risks are detailed in U.S. Pat.
No. 4,923,620 and in U.S. Pat. No. 4,880,548.
In the above described centrifugal method for separating blood into the
three basic fractions, the leucocytes are present in substantial
quantities in both the packed red cells and platelet-rich plasma
fractions. It is now generally accepted that it would be highly desirable
to reduce the leucocyte concentration of these blood components to as low
a level as possible. While there is no firm criterion, it is generally
accepted that many of the undesirable effects of transfusion would be
reduced if the leucocyte content were reduced by a factor of about 100 or
more prior to administration to the patient. This approximates reducing
the average total content of leucocytes in a single unit of PRC or PRP to
less than about 1.times.10.sup.7, and in a unit of PRP or PC to less than
about 1.times.10.sup.6.
Growing evidence suggests that the use of leucocyte depleted platelet
concentrates decreases the incidence of febrile reactions and platelet
refractoriness. Leucocyte depleted blood components are also believed to
have a role in reducing the potential for Graft Versus Host Disease.
Leucocyte depletion of platelet preparations is also believed to diminish,
but not to completely prevent, the transmission of leucocyte associated
viruses such as HIV-1 and CMV.
Platelet preparations contain varying amounts of leucocytes. The level of
leucocyte contamination in unfiltered conventional platelet preparations
of 6 to 10 pooled units is generally at a level of about 5.times.10.sup.8
or greater. Platelet concentrates prepared by the differential
centrifugation of blood components will have varying levels of leucocyte
contamination related to the time and to the magnitude of the force
developed during centrifugation. It has been demonstrated that leucocyte
removal efficiencies of 81 to 85% are sufficient to reduce the incidence
of febrile reactions to platelet transfusions. Several other recent
studies report a reduction in alloimmunization and platelet refractoriness
at levels of leucocyte contamination <1.times.10.sup.7 per unit. For a
single unit of PC, the goal is to reduce the number of leucocytes from
about 7.times.10.sup.7 leucocytes (average leucocyte contamination level
under current practice) to less than about 1.times.10.sup.6 leucocytes.
The existing studies therefore suggest the desirability of at least a two
log (99%) reduction of leucocyte contamination. More recent studies
suggest that a three log (99.9%) or even a four log (99.99%) reduction
would be significantly more beneficial.
An additional desirable criterion is to restrict platelet loss to about 15%
or less of the original platelet concentration. Platelets are notorious
for being "sticky", an expression reflecting the tendency of platelets
suspended in blood plasma to adhere to any non-physiological surface to
which they are exposed. Under many circumstances, they also adhere
strongly to each other.
In any system which depends upon filtration to remove leucocytes from a
platelet suspension, there will be substantial contact between platelets
and the internal surfaces of the filter assembly. The filter assembly must
be such that the platelets have minimal adhesion to, and are not
significantly adversely affected by contact with, the filter assembly's
internal surfaces.
U.S. Pat. No. 4,880,548 provides a convenient and very effective means for
leuco-depleting PC. PC is passed through a fibrous porous medium which
permits recovery of 90% or more of the platelets, which pass through the
medium, while retaining within the medium more than 99.9% of the incident
leucocytes. This system is currently in widespread use at bedside in
hospitals, but, unlike the device of this invention, it is not as well
suited for use in blood banks during the processing of donated whole
blood. The unsuitability stems primarily from additional storage
constraints associated with PC and the methods of administering PC. For
example, platelets in PC are typically suspended in a total volume of only
about 40 to 60 ml of plasma. Contrasted with this, the platelets which are
processed by the devices and methods of this invention are typically
derived from a single unit of whole blood and are suspended as PRP in
about 180 to 240 ml of plasma.
Further, the platelets in PC have been subjected, during two centrifugation
steps, to severe conditions and may not as readily disperse. It has been
suggested that the high forces to which the platelets are subjected as
they reach the bottom of the bag during sedimentation, promote increased
aggregation by particle-to-particle adhesion.
For these and perhaps other reasons, platelets in PC show a much higher
tendency to be retained within the filter during leucocyte depletion
compared with platelets in PRP. Accordingly, a much better recovery is
obtained when platelets are leucocyte-depleted in the form of PRP,
compared with PC; for example, while optimal recovery from PC is about 90
to 95%, recovery from PRP can exceed 99%.
Also, as a consequence of the concentration differences and possibly as a
consequence of the lower degree of aggregation in PRP, the preferred
critical wetting surface tension (CWST) range when filtering PRP is
broader than that for PC.
Devices which have previously been developed in attempts to meet the
above-noted objectives have been based on the use of packed fibers, and
have generally been referred to as filters. However, it would appear that
processes utilizing filtration based on separation by size cannot succeed
for two reasons. First, leucocytes can be larger than about 15 .mu.m
(e.g., granulocytes and macrocytes) to as small as 5 to 7 .mu.m (e.g.,
lymphocytes). Together, granulocytes and lymphocytes represent the major
proportion of all of the leucocytes in normal blood. Red blood cells are
about 7 .mu.m in diameter, i.e., they are about the same size as
lymphocytes, one of the two major classes of leucocytes which must be
removed. Secondly, all of these cells deform so that they are able to pass
through much smaller openings than their normal size. Accordingly, it has
been widely accepted that removal of leucocytes is accomplished mainly by
adsorption on the internal surfaces of porous media, rather than by
filtration.
The separation of the various blood components using centrifugation is
attended by a number of problems. First, in the separation of
platelet-rich plasma from PRC, e.g., step 3 above, it is difficult to
efficiently obtain the maximum yield of platelets while preventing red
cells from entering the plasma. Secondly, when PRP is expressed, it is
difficult to efficiently recover the more desirable younger platelets
located near or in the PRC/PRP interface.
BRIEF SUMMARY OF THE INVENTION
In the methods of this invention, leucocyte depletion is preferably
accomplished at the time the blood is processed. During the separation of
PRP from PRC, the process may be enhanced by interposing a red cell
barrier medium immediately downstream of the blood collection bag. Thus,
the supernatant PRP passes through a porous medium until red cells block
the medium. The platelet-containing solution such as PRP may be
subsequently centrifuged to obtain a supernatant leucocyte-depleted plasma
layer and a sediment leucocyte-depleted PC layer. The method and apparatus
of the present invention permit the recovery of an increased amount of
more desirable platelets and of plasma more efficiently in comparision to
conventional blood processing practices.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 is a cross section view of an embodiment of a red cell barrier
filter assembly, taken along A--A of FIG. 2a.
FIG. 2a is a top view of an embodiment of a red cell barrier filter
assembly according to the invention.
FIG. 2b is a bottom view of an embodiment of a red cell barrier filter
assembly according to the invention.
FIG. 3 is an embodiment of a biological fluid processing system according
to the invention, whereby a red cell barrier filter assembly is interposed
between a collection container and a satellite bag.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
The present invention involves a method for harvesting an increased amount
of platelets from a platelet-containing solution, particularly PRP, which
comprises separating a red cell containing biological fluid such as whole
blood into a red cell-containing sediment layer and a non-red cell
containing supernatant layer, and passing the supernatant layer of the
separated fluid through a filter until red cells block the filter. An
increased amount of platelets and/or plasma may then be recovered. The
present invention also involves an apparatus which permits the increased
recovery of platelets comprising a porous medium which passes platelets
and/or plasma therethrough, but blocks the passage of red cells. The
present invention also involves a system for harvesting an increased
amount of platelets and/or plasma which comprises a first container in
fluid communication with second container, and, interposed between the
first container and the second container, a red cell barrier medium.
An exemplary biological fluid red cell barrier filter assembly is shown in
FIGS. 1 and 2. A red cell barrier filter assembly may include a housing 1
having an inlet 2 and an outlet 3 and defining a liquid flow path between
the inlet and the outlet. A red cell barrier medium 4, preferably
positioned inside the housing across the liquid flow path, includes a
porous medium which permits a platelet-containing solution such as PRP to
pass therethrough, but blocks the passage of red cells. In a preferred
embodiment, flow through the filter assembly is stopped automatically when
red cells contact the red cell barrier medium.
While the red cell barrier medium can be produced from any suitable
material compatible with a biological fluid such as blood, practical
considerations dictate that consideration be given first to the use of
commercially available materials. The porous medium of this invention may
be formed, for example, from any synthetic polymer capable of forming
fibers and cf serving as a substrate for grafting. Preferably, the polymer
should be capable of reacting with at least one ethylenically unsaturated
monomer under the influence of ionizing radiation without the matrix being
significantly or excessively adversely affected by the radiation. Suitable
polymers for use as the substrate include, but are not limited to,
polyolefins, polyesters, polyamides, polysulfones, acrylics,
polyacrylonitriles, polyaramides, polyarylene oxides and sulfides, and
polymers and copolymers made from halogenated olefins and unsaturated
nitriles. Examples include, but are not limited to, polyvinylidene
fluoride, polyethylene, polypropylene, cellulose acetate, and Nylon 6 and
66. Preferred polymers are polyolefins, polyesters, and polyamides. The
most preferred polymer is polybutylene terephthalate (PBT).
Although the fibers of the porous medium may remain untreated, they are
preferably treated to make them even more effective. For example, the
fibers may be surface modified to increase the critical wetting surface
tension (CWST) of the fibers.
Surface characteristics of a fiber can be modified by a number of methods,
for example, by chemical reaction including wet or dry oxidation, by
coating the surface by depositing a polymer thereon, by grafting reactions
which are activated by exposure to an energy source such as gas plasma,
heat, a Van der Graff generator, ultraviolet light, or to various other
forms of radiation, or by surface etching or deposition using a gas plasma
treatment. The preferred method is a grafting reaction using
gamma-radiation, for example, from a cobalt source.
In a preferred form of the porous medium of the subject invention, the
fibers of which the filter element is composed may be modified by grafting
thereon a mixture of two monomers, one containing hydroxyl groups and
another containing anionic groups, such as carboxyl groups, with the
hydroxyl groups present in larger numbers. As described in U.S. Pat. No.
4,880,548, the filter media of this invention are preferably surface
modified using a mixture comprising hydroxyl-terminated and
carboxyl-terminated monomers. In a preferred form of this invention, the
monomers are respectively hydroxyethyl methacrylate (HEMA) and methacrylic
acid (MAA), and the monomer ratios (carboxyl:hydroxyl) are preferably in
the range of about 0.01:1 to about 0.5:1, and more preferably in the range
of about 0.05:1 to about 0.35:1. A preferred monomer ratio is one which
produces a desired zeta potential at the pH of plasma (7.3) of about -3 to
about -30 millivolts, a more preferred ratio produces a zeta potential of
about -7 to about -20 millivolts, and a still more preferred ratio
produces a zeta potential of about -10 to about -14 millivolts.
Radiation grafting, when carried out under appropriate conditions, has the
advantage of considerable flexibility in the choice of reactants,
surfaces, and in the methods for activating the required reaction.
Gamma-radiation grafting is particularly preferable because the products
are very stable and have undetectably low aqueous extractable levels.
Furthermore, the ability to prepare synthetic organic fibrous media having
a CWST within a desired range is more readily accomplished using a gamma
radiation grafting technique.
An exemplary radiation grafting technique may employ at least one of a
variety of monomers each comprising an ethylene or acrylic moiety and a
second group, which can be selected from hydrophilic groups (e.g., --COOH,
or --OH). Grafting of the fibrous medium may also be accomplished by
compounds containing an ethylenically unsaturated group, such as an
acrylic moiety, combined with a hydroxyl group, preferably monomers such
as HEMA or acrylic acid. The compounds containing an ethylenically
unsaturated group may be combined with a second monomer such as MAA. Use
of HEMA as the monomer contributes to a very high CWST. Analogues with
similar functional characteristics may also be used to modify the surface
characteristics of fibers.
The number of carboxyl groups per unit of surface area appears to have an
important effect on the adhesion of platelets to fiber surfaces. This
effect is reflected in the proportion of platelets recovered in the filter
effluent as a fraction of the number present prior to filtration. Platelet
recovery typically peaks at the optimum proportion of MAA. The number of
carboxyl groups per unit of fiber surface is, over the range of interest
of this invention, thought to be close to proportional to the amount of
MAA in the monomeric grafting solution.
The CWST of the porous media made with the PBT fibers typically have a CWST
as formed of about 50 to about 54 dynes/cm, and most or all other fibers
which may be used have a CWST below about 55 dynes/cm. Surface grafting
using the monomers noted above causes the CWST of the fibers to increase,
the exact value obtained being dependent on the ratio of the two monomers.
A preferred range for the CWST of the devices of this invention is greater
than about 70 dynes/cm, typically from about 70 dynes/cm to about 115
dynes/cm a more preferred range is about 90 to about 100 dynes/cm and a
still more preferred range is about 93 to about 97 dynes/cm, these ranges
being obtained by varying the ratio of carboxyl-terminated and
hydroxyl-terminated monomers.
As disclosed in U.S. Pat. No. 4,880,548, the CWST of a porous medium may be
determined by individually applying to its surface a series of liquids
with surface tensions varying by 2 to 4 dynes/cm and observing the
absorption or non-absorption of each liquid over time. The CWST of a
porous medium, in units of dynes/cm, is defined as the mean value of the
surface tension of the liquid which is absorbed and that of the liquid of
neighboring surface tension which is not absorbed within a predetermined
amount of time. The absorbed and non-absorbed values depend principally on
the surface characteristics of the material from which the porous medium
is made and secondarily on the pore size characteristics of the porous
medium.
Liquids with surface tensions lower than the CWST of a porous medium will
spontaneously wet the medium on contact and, if the medium has through
holes, will flow through it readily. Liquids with surface tensions higher
than the CWST of the porous medium may not flow at all at low differential
pressures and may do so unevenly at sufficiently high differential
pressures to force the liquid through the porous medium. In order to
achieve adequate priming of a fibrous medium with a liquid such as blood,
the fibrous medium preferably has a CWST in the range of about 53 dynes/cm
or higher.
A red cell barrier filter assembly produced in accordance with the present
invention and suitable for passing about one unit of PRP preferably has a
fiber surface area of about 0.04 to about 3.0 M.sup.2, more preferably
about 0.06 to about 2.0 M.sup.2. A preferred range for the filter element
flow area is about 3 to about 8 cm.sup.2, more preferably about 4 to about
6 cm.sup.2. A preferred range for the relative voids volume is about 71%
to about 83% (corresponding for PBT fibers to a density of about 0.23 to
about 0.40 g/cc), more preferably about 73% to about 80% (about 0.27 to
about 0.37 g/cc). Because of its small size, a preferred filter in
accordance with the present invention retains internally only about 0.5 to
1 cc of PRP, representing less than a 0.5% loss of platelets.
In another embodiment of the invention, the fiber may be surface modified
in the same manner as noted above, but the fiber surface area of the
element is increased while, at the same time, the density of the filter
element is somewhat reduced. In this way, the automatic blockage of flow
on contact by red cells is combined with higher efficiency of leucocyte
depletion.
A preferred range of fiber surface area for this embodiment of the
invention is from about 0.3 to about 2.0 M.sup.2, and a more preferred
range is from about 0.35 to about 0.6 M.sup.2. The upper limits of fiber
surface area reflect the desire to accomplish the filtration in a
relatively short time period, and may be increased if longer filtration
times are acceptable. A preferred voids volume of a porous medium of this
embodiment is in the range of about 71% to about 83% (i.e., if PBT fiber
is used, corresponding to a density of the filter element in the range of
about 0.23 g/cc to about 0.40 g/cc), and more preferably about 75% to
about 80% (for PBT, about 0.28 g/cc to about 0.35 g/cc). A preferred
filter element flow area is from about 2.5 to about 10 cm.sup.2, and a
more preferred area is from about 3 to about 6 cm.sup.2. The upper limits
of the filter element flow area reflect the desire to accomplish the
filtration in a relatively short time period, and may be increased if
longer filtration times are acceptable. Leucocyte depletion efficiencies
in excess of about 99.9 to about 99.99%, which corresponds to an average
residual leucocyte content per unit of less than about
0.005.times.10.sup.7, can be obtained.
Although the porous medium of the present invention may have a
substantially uniform density, the porous medium of a preferred embodiment
of the present invention is of a construction such that an upstream
portion of the porous medium is of generally lower density than a
downstream portion of the filter. For example, the density of the porous
medium may vary in a continuous or stepwise manner while maintaining an
average density range suitable for blocking red cells. An exemplary porous
medium may include a density range in the upstream portion from about 0.1
g/cc to about 0.23 g/cc, and a density range in the downstream portion
from about 0.23 g/cc to about 0.40 g/cc. In another embodiment of the
invention, the porous medium may include two or more layers, preferably of
different or varying density. An exemplary zoned or layered medium is
illustrated in FIG. 1; using PBT as the fiber upstream layer 5 may include
a density range from about 0.1 g/cc to about 0.2 g/cc, middle layer 6 may
include a density range from about 0.20 g/cc to about 0.25 g/cc, and
downstream layer 7 may include a density range from about 0.23 g/cc to
about 0.40 g/cc.
Included within the scope of the present invention are the use of other
density valves, in a particular zone or layer as well as throughout the
porous medium. These alternative density ranges may be chosen based on
achieving a desired result, in addition to blocking red cells, e.g., the
flow rate, the type of fiber used, the amount of leucocytes removed, as
well as other considerations.
The porous medium may act as an automatic "valve" by instantly stopping the
flow of the supernatant layer of the centrifuged whole blood, which
supernatant layer will typically be a platelet-rich solution such as PRP,
when red cells from the sediment layer, typically a red cell containing
solution such as PRC, contact the porous medium. The mechanism of this
valve-like action may reflect aggregation of the red cells concentrated at
the PRP/PRC transition zone (buffy coat) as they reach the medium's
surface, forming a barrier which prevents or blocks further flow of the
supernatant layer through the porous medium. Aggregation of red blood
cells on contact with the porous medium appears to be related to the CWST
and/or to other less understood surface characteristics of the fibers.
This theory for the proposed mechanism is supported by the existence of
filters capable of highly efficient leucocyte depletion of human red blood
cell suspensions and which have pore sizes as small as 0.5 .mu.m, through
which red cells pass freely and completely with no clogging, with applied
pressure of the same magnitude as that used in the present invention. On
the other hand, the filters of the present invention, which typically have
pore diameters larger than about 0.5 .mu.m, abruptly stop the flow of red
blood cells when the porous medium is contacted by the red cells.
Housings for the filter assembly to be used in conjunction with the present
invention can be fabricated from any suitably impervious material,
including an impervious thermoplastic material. For example, the housing
may preferably be fabricated by injection molding from a transparent or
translucent polymer, such as an acrylic, polystyrene, or polycarbonate
resin. Not only is such a housing easily and economically fabricated, but
it also allows observation of the passage of the fluid through the
housing.
Any housing of suitable shape, preferably providing an inlet and an outlet,
may be employed. The housing may include an arrangement of one or more
channels, grooves, conduits, passages, ribs, or the like, which may be
serpentine, parallel, curved, circular, or a variety of other
configurations. An exemplary embodiment is shown in FIGS. 2A and 2B,
illustrating a circular housing 1 having an inlet 2 and an outlet 3. A
preferred embodiment of the invention includes one or more ribs 8 on the
upstream side of the housing and at least one channel or groove on the
downstream side of the housing. In a most preferred embodiment of the
invention, the housing 1 includes a series of concentric grooves or
channels 9a and radial grooves or channels 9b which provide fluid
communication with the outlet 3.
The housing into which the porous medium is placed may be sealed or
interference fit, and is designed to achieve practical and economic
construction, convenience of use, rapid priming, and efficient air
clearance.
The porous components of devices made in accordance with the invention are
preferably preformed prior to assembly to controlled dimension and pore
diameter in order to form an integral self-contained element.
Preforming eliminates the pressure on the inlet and outlet faces of the
container which are inherent in a packed fiber system. Pre-forming the
porous element typically leads to devices having longer service life,
coupled with at least equal and usually better leucocyte removal
efficiency, equal or better platelet recovery, and less hold up of fluid,
when compared to devices that use fibers or fibrous webs packed into a
housing at assembly.
Furthermore, pre-forming enhances the proper positioning of the porous
medium in the housing. The lateral dimensions of the porous element are
typically larger than the corresponding dimensions of the housing into
which they are assembled. For example, if the porous medium is in disc
form, the outside diameter of the pre-formed medium is made about 1%
larger than the housing inside diameter. This provides very effective
sealing by an interference fit with no loss of effective area of the
porous medium, and contributes further towards minimization of the fluid
hold-up volume of the assembly. In accordance with the invention,
assembling the porous medium in the housing using an interference fit seal
is preferred. However, edge compression about the periphery, a compression
seal, or other means of positioning the porous medium in the housing may
be used.
Included within the scope of the present invention is the inclusion of the
red cell barrier medium or filter assembly in biological fluid processing
systems, preferably closed, sterile systems, having a wide variety of
components, such as one or more biological fluid collection bags; one or
more satellite bags; gas or air inlets and outlets; and/or one or more
connectors, such as SCD connectors.
An exemplary biological fluid collection and processing system is shown in
FIG. 3. The biological fluid processing system is generally denoted as 10.
It may comprise a first container or collection bag 11; a needle or
cannula 50 adapted to be inserted into the donor; a red cell barrier
filter assembly 12; a first leucocyte depletion assembly 13 (optional); a
second container (first satellite bag) 41, typically for receiving a
platelet-rich solution or suspension 31; an optional fourth container
(third satellite bag) 42, typically for receiving platelet concentrate; a
second leucocyte depletion assembly 17; and a third container (second
satellite bag) 18, typically for receiving a red cell containing solution
or suspension 32. Each of the assemblies or containers may be in fluid
communication through tubing, preferably flexible tubing, 20, 21, 25, 26,
27 or 28. The first leucocyte depletion assembly preferably includes a
porous medium for passing PRP; the second leucocyte depletion assembly
preferably includes a porous medium suitable for passing PRC. A seal,
valve, clamp, or transfer leg closure (not illustrated) may also be
positioned in or on the tubing or in the collection and/or satellite bags.
The seal (or seals) is opened when fluid is to be transferred between
bags.
The invention also involves a method for processing a biological fluid
containing red blood cells comprising collecting whole blood in a
container; forming a supernatant layer and a sediment layer, typically by
differential sedimentation such as centrifugation; and passing the
supernatant layer through a porous medium, the porous medium comprising a
red cell barrier medium or a combined leucocyte depletion red cell barrier
medium. The supernatant layer passes through the porous medium until red
cells contact the porous medium, at which point flow through the medium
stops automatically.
In general, donated whole blood is processed as soon as practicable in
order to more effectively reduce or eliminate contaminating factors,
including but not limited to leucocytes and microaggregates. In accordance
with the subject invention, leucocyte depletion may be accomplished during
the initial processing of the whole blood, which in United States practice
is generally within 8 hours of collection from the donor. After the
cellular component of whole blood, i.e., red cells, have separated, the
liquid portion, i.e. supernatant PRP, is expressed from the blood
collection bag into a first satellite bag through one or more porous media
which diminish the amount of leucocytes and/or block red cells.
In general, using the Figures for reference, the biological fluid (e.g.,
donor's whole blood) is received directly into the collection bag 11. The
collection bag 11, with or without the other elements of the system, may
then be centrifuged in order to separate the biological fluid into a
supernatant layer, typically a platelet-containing solution such as PRP,
and a sediment layer, typically a red cell solution such as PRC. The
biological fluid may be expressed from the collection bag as separate
supernatant and sediment layers, respectively. There may be a clamp or the
like on or in the bag or tubing to prevent the flow of the supernatant
layer from entering the wrong conduit.
Movement of the biological fluid through the system is effected by
maintaining a pressure differential between the collection bag and the
destination of the biological fluid (e.g., a container such as a satellite
bag). Exemplary means of establishing this pressure differential may be by
expressor, gravity head, applying pressure to the collection bag (e.g., by
hand or with a pressure cuff), or by placing the other container (e.g.,
satellite bag) in a chamber (e.g., a vacuum chamber) which establishes a
pressure differential between the collection bag and the other container.
Also included within the scope of the invention may be expressors which
generate substantially equal pressure over the entire collection bag.
As the biological fluid passes from one bag to the next, it may pass
through at least one porous medium. Typically, if the biological fluid is
the supernatant layer (e.g., PRP), it may pass from the collection bag
through one or more devices or assemblies comprising one or more porous
media--a leucocyte-depletion medium, a red cell barrier medium, a porous
medium which combines the red cell barrier with leucocyte depletion in one
porous medium, or a leucocyte depletion medium and a red cell barrier
medium in series. The supernatant layer is expressed from the first
container 11 until flow is stopped. Additional processing, if desired, may
occur downstream of the red cell barrier medium, either connected to the
system or after being separated from the system.
In accordance with an additional embodiment of the invention, a method is
provided whereby the recovery of various biological fluids is maximized.
Recovery of an increased amount of PRP in and of itself may increase the
amount of platelets recovered. Furthermore, recovering a greater amount of
the platelets located in or near the PRP/PRC interface may increase the
recovery of the more useful and/or more desirable younger platelets.
The advantages to be gained by the use of the methods and devices of the
invention include the following:
(a) The PC derived from the PRP is substantially free of red cells, and may
include a higher proportion of younger platelets.
(b) The operator needs only to start the flow of platelet-rich solution,
which will continue to flow into the first satellite bag until red cells
contact the filter surface, at which point flow stops automatically. This
eliminates the need for a skilled operator to estimate when to stop flow
and decreases the possibility of red cell contamination.
(c) The volume of plasma and PC recovered from the blood collection bag
during the extraction operation may be increased by about 5% or more when
compared with very competent manual operation, and the concentration of
platelets recovered may be increased by about 15% to about 30% or more.
(d) About 90% or greater of the platelets in whole blood are recovered.
(e) Labor input is reduced, as monitoring of the interface during
decantation is not required.
(f) Freshly donated blood contains platelets varying in age from newly
formed to nine days or more (platelet half-life in vivo is about nine
days). Newly formed platelets are larger and are generally believed to be
more active. Because the younger platelets are larger, they tend to
sediment faster during centrifugation and, consequently, are present in
larger numbers in the PRP nearest to the red cell interface. Measurements
have shown that the concentration of platelets in the 10% of the PRP
volume nearest the interface is about twice that in the uppermost 10% of
PRP. Taking this into account, the total number of platelets recovered may
be increased by about 4 to 10%.
______________________________________
Incremental number
of platelets, %
______________________________________
Due to increased volume of PRP
2 to 5
Due to the higher concentration
2 to 5
of platelets in the incremental
volume of PRP
Total 4 to 10%
______________________________________
(g) The larger proportion of younger platelets in the PC administered to
the patient means that their life within the patient after administration
will be longer and that the platelets will be more active, compared with
current blood bank practice.
(h) The yield of plasma, a component of value comparable with that of PRC
and PC, may also increased by about 2 to about 5%.
(i) Insofar as the plasma yield is increased, the plasma content of the PRC
is decreased. This is advantageous because the MHC (major
histocompatibility complex) contained in the plasma is responsible for the
occurrence of Urticaria (hives) in a proportion of transfusion recipients
transfused with PRC.
Definitions: The following definitions are used in reference to the
invention:
A) Blood Product or Biological Fluid: anti-coagulated whole blood (AWB);
packed red cells obtained from AWB; platelet-rich plasma (PRP) obtained
from AWB; platelet concentrate (PC) obtained from AWB or PRP; plasma
obtained from AWB or PRP; red cells separated from plasma and resuspended
in physiological fluid; and platelets separated from plasma and
resuspended in physiological fluid. Blood product or biological fluid also
includes any treated or untreated fluid associated with living organisms,
particularly blood, including whole blood, warm or cold blood, and stored
or fresh blood; treated blood, such as blood diluted with a physiological
solution, including but not limited to saline, nutrient, and/or
anticoagulant solutions; one or more blood components, such as platelet
concentrate (PC), platelet-rich plasma (PRP), platelet-free plasma,
platelet-poor plasma, plasma, packed red cells (PRC), or buffy coat;
analogous blood products derived from blood or a blood component or
derived from bone marrow. The biological fluid may include leucocytes, or
may be treated to remove leucocytes. As used herein, blood component or
product refers to the components described above, and to similar blood
products obtained by other means and with similar properties. In
accordance with the invention, each of these blood products is processed
in the manner described herein.
B) Unit of Whole Blood: blood banks in the United States commonly draw
about 450 milliliters (ml) of blood from the donor into a bag which
contains an anticoagulant to prevent the blood from clotting. However, the
amount drawn differs from patient to patient and donation to donation.
Herein the quantity drawn during such a donation is defined as a unit of
whole blood.
C) Unit of Packed Red Cells (PRC), Platelet-rich Plasma (PRP), or Platelet
Concentrate (PC): As used herein, a "unit" is defined by the United
States' practice, and a unit of PRC, PRP, PC, or of red cells or platelets
in physiological fluid or plasma, is the quantity derived from one unit of
whole blood. Typically, the volume of a unit varies. For example, the
volume of a unit of PRC varies considerably dependent on the hematocrit
(percent by volume of red cells) of the drawn whole blood, which is
usually in the range of about 37% to about 54%. The concomitant hematocrit
of PRC, which varies over the range from about 50 to over 80%, depends in
part on whether the yield of one or another blood product is to be
minimized. Most PRC units are in the range of about 170 to about 350 ml,
but variation below and above these figures is not uncommon.
D) Porous medium: refers to the porous medium through which one or more
blood components pass. The porous medium refers generically to any one of
the media which deplete leucocytes from the non-PRC blood components,
i.e., from PRP or from PC and/or which block the passage of red cells
while allowing the passage of platelets and plasma.
The porous medium for use with a platelet-rich solution such as PRP may be
formed from any natural or synthetic fiber or other porous material
compatible with blood. Preferably, the CWST and zeta potential of the
porous medium are within certain ranges, as disclosed above and as
dictated by its intended use. For example, the CWST of a PRP porous medium
is typically above about 70 dynes/cm.
The porous medium may be configured as a flat sheet, a composite of two or
more layers, a corrugated sheet, a web, a fibrous mat, a depth filter or a
membrane, although it is not intended that the invention should be limited
thereby.
E) Voids volume is the total volume of all of the pores within a porous
medium. Voids volume is expressed hereinafter as a percentage of the
apparent volume of the porous medium.
F) Conversion of density when using fibers other than PBT: In the preceding
exposition the term density has been used, and the density values quoted
for the filter element have been based on the use of PBT fibers. Other
fibers which differ in density from the PBT may be used, as noted above,
providing that their surfaces have, or have been modified to have, the
characteristics noted above, e.g., a CWST of greater than 70 dynes/cm. In
accordance with the invention, to use an alternate fiber of different
density, the density of an element made using an alternate fiber (i.e.,
the PBT equivalent density) may be calculated as follows:
Denoting V as a percentage of the voids volume relative to the apparent
volume of the PBT element [i.e., V=(volume of voids/volume of
element).times.100], the objective is to calculate the element density of
an alternate fiber element which will have a relative voids volume
percentage equal to V.
If F is the density of the alternate fiber and 1.38 g/cc is taken as the
density of PBT fiber, and M.sub.1 is the element density of the PBT
element and M.sub.2 is the density required for an element with equivalent
performance, then voids volume V of the PBT fiber element is
V=(1-M.sub.1 /1.38).times.100
and the density required for the element made using the alternate fiber is
M.sub.2 =F (1-V/100).
The more preferred fiber diameter range for the practice of this invention
is about 2 to 3 .mu.m, the diameter being defined in terms of surface
area, as described in U.S. Pat. No. 4,880,548. This range is preferred
because much above this range, the dimensions of the elements and
consequently the liquid hold-up volumes of the filters become
significantly larger; below this range, the filter elements become
relatively less coherent and are more easily compressed. For example, an
element made using less than 2 .mu.m polypropylene fibers would be
compressed by the pressure developed by the plasma extractor, which can be
as high as 300 mm of Hg.
Pore diameters of filter elements in accordance with the invention can be
determined using the modified OSU F2 method as described in U.S. Pat. No.
4,925,572. Filter assemblies with good efficiency and recovery can be made
using large pore diameters, but such filter assemblies typically retain a
higher proportion of platelets. A filter assembly having a pore diameter
of about 15 .mu.m to 30 .mu.m or higher may allow some red cells and
leucocytes to pass, thereby reducing platelet recovery efficiency.
Therefore, it is preferred that the pore diameter not exceed 15 .mu.m,
more preferably, less than about 10 .mu.m. The most preferred pore
diameter range is less than about 6 .mu.m.
G) In accordance with the invention, a useful technique for the measurement
of fiber surface area, for example by nitrogen gas adsorption, is that
developed by Brunauer, Emmet, and Teller in the 1930's, often referred to
as the "BET" measurement. Using PBT as an example, the surface area of
meltblown webs can be used to calculate average fiber diameter:
##EQU1##
where L=total length in cm of 1 gram of fiber, d=average fiber diameter in
centimeters, and
A.sub.f =fiber surface area in cm.sup.2 /g.
If the units of d are micrometers, the units of A.sub.f become M.sup.2 /g
(square meters/gram), which will be used hereinafter. For fibers other
than PBT, substitute the density for 1.38.
H) General procedure for measuring zeta potential: Zeta potential was
measured using a sample cut from a 1/2 inch thick stack of webs.
The zeta potential was measured by placing the sample in an acrylic filter
holder which held the sample snugly between two platinum wire screens
100.times.100 mesh (i.e., 100 wires in each direction per inch). The
meshes were connected, using copper wire, to the terminals of a Triplett
Corporation model 3360 Volt-Ohm Meter, the mesh on the upstream side of
the sample being connected to the positive terminal of the meter. A
pH-buffered solution was flowed through the sample using a differential
pressure of 45 inches of water column across the filter holder and the
effluent was collected. For measurements at pH 7, a buffered solution was
made by adding 6 ml pH 7 buffer (Fisher Scientific Co. catalog number
SB108-500) and 5 ml pH 7.4 buffer (Fisher Scientific Co. catalog number
SB110-500) to 1 liter pyrogen-free deionized water. For measurements at pH
9, a buffered solution was made by adding 6 ml pH 9 buffer (Fisher
Scientific Co. catalog number SB114-500) and 2 ml pH 10 buffer (Fisher
Scientific Co. catalog number SB116-500) to 1 liter pyrogen-free deionized
water. The electrical potential across the filter holder was measured
during flow (it required about 30 seconds of flow for the potential to
stabilize) and was corrected for cell polarization by subtracting from it
the electrical potential measured when flow was stopped. During the period
of flow the pH of the liquid was measured using a Cole-Parmer model
J-5994-10 pH meter fitted with an in-line model J-5993-90 pH probe. The
conductivity of the liquid was measured using a Cole-Parmer model
J-1481-60 conductivity meter fitted with a model J-1481-66 conductivity
flow cell. Then the polarity of the volt meter was reversed, and the
effluent was flowed backwards through the filter holder using a
differential pressure of 45 inches of water column. As in the first
instance the electrical potential measured during flow was corrected for
cell polarization by subtracting from it the electrical potential measured
after flow was stopped. The average of the two corrected potentials was
taken as the streaming potential.
The zeta potential of the medium was derived from the streaming potential
using the following relationship (J. T. Davis et al., Interfacial
Phenomena, Academic Press, New York, 1963):
##EQU2##
where .sup.n is the viscosity of the flowing solution, D is its dielectric
constant, .lambda. is its conductivity, E.sub.s is the streaming potential
and P is the pressure drop across the sample during the period of flow. In
these tests the quantity 4 .pi..sup.n /DP was equal to 0.800.
EXAMPLES
Each of the examples was run using the following basic procedure to process
and test a bag of donated blood. The blood collection set was constituted
as shown in FIG. 3. Bag 11, into which anticoagulant had been placed, was
used to collect one unit of about 450 cc of blood from a human volunteer.
Bag 11 along with its two satellite bags 18, 41 was then centrifuged for 5
minutes at 2280.times.gravity, causing the red cells to sediment into the
lower parts of the collection bag and leave a transparent, yellowish layer
of red cell-free plasma in the upper part of the collection bag. This bag
was then transferred, with care not to disturb its contents, to a plasma
extractor. With tube 20 clamped adjacent to bag 11 to prevent flow, tube
20 was cut and red cell barrier filter assembly 12 and/or leucocyte
depletion filter assembly 13 were inserted at the position as shown in
FIG. 3. With the plasma extractor applying sufficient force to the bag to
generate a pressure of about 200 to 300 millimeters of mercury within the
bag, the clamp on tube 20 was removed, allowing the supernatant liquid to
flow through the filter assemblies 12 and/or 13 into bag 41 which had been
placed on a weight scale. One of several skilled operators was instructed
to signal when, in normal blood bank practice, flow would have been
manually shut off. For examples 1 and 2, which were in accordance with an
embodiment of the invention having a PRP leucocyte depletion filter
assembly 13, tube 20 was at the signal promptly shut-off, the weight of
PRP collected was recorded, and the contents of the bag analyzed, with
results recorded in Table I.
For examples 3-8 and 9-10, the weight of the PRP bag 41 was recorded at the
signal, i.e., the precise moment when flow would in normal blood bank
practice have been shut off, while flow was allowed to continue until the
red cell layer reached red cell barrier filter assembly 12, at which time
flow automatically and abruptly stopped, and the weight of PRP collected
was recorded. The results for examples 3-8 are shown in Table II, and for
examples 9 and 10 in Table III.
In each of the ten examples, the resulting PRP was visually free of red
cells, and weights of the PRP were converted to volume by dividing by the
density of plasma (1.04 g/cc). The data on residual leucocyte content of
the PC derived from the filtered PRP are reported in Tables II and III as
multiples of 10.sup.7 (i.e., .times.10.sup.7), which can be conveniently
compared with a target criterion of fewer than about 1.times.10.sup.7
leucocytes per unit, which is a level believed adequate to significantly
reduce alloimmunization in patients receiving platelet transfusions.
The widely used melt blowing process for making fibrous plastic webs is a
convenient, economical, and effective means for manufacturing fibrous webs
with fiber diameter in the 1-4 .mu.m range. It is characteristic of this
process that the quality of melt blown webs is optimal when the web weight
is maintained in a preferred range of about 0.0005 to about 0.01
g/cm.sup.2, and more preferably between about 0.0005 and about 0.007
g/cm.sup.2. For this reason, the webs used to form the examples of this
invention were, wherever necessary, formed by laying up two or more layers
of web of weight about 0.006 g/cm.sup.2, and then hot compressing these to
form an integral filter element.
EXAMPLES 1-2
PRP leucocyte depletion filter assemblies were prepared in the manner
described in the specification. The filter elements of these devices were
preformed from 2.6 .mu.m average diameter PBT fibers, which had been
surface modified in the manner as described above and as taught in U.S.
Pat. No. 4,880,548 using a mixture of hydroxyethyl methacrylate and
methacrylic acid in a monomer ratio of 0.35:1 to obtain a CWST of 95
dynes/cm and a zeta potential of -11.4 millivolts. Filter element
effective diameter was 4.74 cm, presenting a filter area of 17.6 cm.sup.2,
thickness was 0.15 cm, voids volume was 83% (density=0.23 g/cc), and fiber
surface area was 0.69 M.sup.2. The volume of PRP held up within the filter
housing was 2.5 cc, representing a loss of PRP due to hold-up of about 1%.
The results, obtained using the operating procedure described earlier in
this section, are shown in Table I.
TABLE I
______________________________________
Leucocyte Depletion Efficiency of the First Variation
Leucocyte
Volume of Leucocyte content of
removal
Example
PRP passed,
PC after filtration
efficiency,**
Number cc (per unit)* %
______________________________________
1 237 <.006 .times. 10.sup.7
>99.9%
2 206 <.006 .times. 10.sup.7
>99.9%
______________________________________
*Total leucocyte count in the PC after centrifuging the filtered PRP to
obtain the PC.
**Assumes that the leucocyte content of the PRP prior to filtration
conformed to an average value of 5 .times. 10.sup.7 per unit.
EXAMPLES 3-8
Red cell barrier filter assemblies were prepared in the manner described in
the specification. The filter elements of these devices were preformed
from 2.6 .mu.m average diameter PBT fibers, which had been surface
modified in the manner as described above and as taught in U.S. Pat. No.
4,880,548 using hydroxyethyl methacrylate and methacrylic acid in a
monomer ratio of 0.35:1 to obtain a CWST of 95 dynes/cm and a zeta
potential of -11.4 millivolts. The filter element's effective diameter was
2.31 cm, presenting a filter area of 4.2 cm.sup.2, thickness was 0.051 cm,
voids volume was 75% (density, 0.34 g/cc), and fiber surface area was 0.08
m.sup.2.
The volume of PRP held up within the filter housing was <0.4 cc,
representing a loss of PRP due to hold-up of less than 0.2%. In each test,
flow stopped abruptly as red cells reached the upstream surface of the
filter element, and there was no visible evidence of red cells or
hemoglobin downstream. The results obtained, using the operating procedure
described earlier in this section for the second variation, are shown in
Table II.
TABLE II
______________________________________
3 5
2 Volume of 4 Leucocyte
Estimated PRP obtained
Incre-
content
volume/PRP using the mental
after filtra-
1 using normal
procedure vol- tion (per
Example
blood bank of invention,
ume, unit of PC* .times.
Number practice, ml
ml percent
10.sup.7
______________________________________
3 175.2 178.8 2.0 1.0
4 212.9 218.8 2.7 1.7
5 221.1 225.7 2.0 0.5
6 185.9 191.4 2.9 0.2
7 257.2 263.2 2.3 <0.1
8 196.6 200.7 2.1 0.1
______________________________________
*Total leucocyte count in the PC after centrifuging the filtered PRP to
obtain PC.
EXAMPLES 9-10
Combined PRP leucocyte depletion/red cell barrier filter assemblies were
prepared in the manner described in the specification i.e., the
combination of an automatic shut-off valve and a high efficiency filter,
both included in a single filter. The filter elements of these devices
were preformed from 2.6 .mu.m average diameter PBT fibers, which had been
surface modified in the manner as described above and as taught in U.S.
Pat. No. 4,880,548 using a mixture of hydroxyethyl methacrylate and
methacrylic acid in a monomer ratio of 0.35:1 to obtain a CWST of 95
dynes/cm and a zeta potential of -11.4 millivolts at the pH of plasma
(7.3). The filter element effective diameter was 2.31 cm presenting a
filter area of 4.2 cm.sup.2 thickness was 0.305 cm, density was 0.31 g/cc
(voids volume=77.5%), and fiber surface area was 0.46 M.sup.2. The volume
of PRP held up within the filter housing was 1.3 cc, representing a loss
of PRP due to hold up within the filter of about 0.5%. In each case, flow
stopped abruptly as red cells reached the upstream surface of the filter
element, and there was no visible evidence of red cells or hemoglobin
downstream. The results obtained, using the operating procedure described
earlier in this section are shown in Table III.
TABLE III
__________________________________________________________________________
Incremental Volume and Leucocyte Depletion Efficiency of the Third
Variation
Volume of PRP Leucocyte content
Estimated volume/PRP
obtained using after filtration
Leucocyte
using normal
the procedure of
Incremental
(per unit)
removal
Example Number
blood bank practice, ml
invention, ml
volume, %
of PC* .times. 10.sup.7
efficiency**
__________________________________________________________________________
9 251 256 2 11 <.004 >99.9%
10 212 216 1.9 .005 >99.9%
__________________________________________________________________________
*Total leucocyte count in the PC after centrifuging the filtered PRP to
obtain PC.
**Assumes that the leucocyte content of the PRP prior to filtration
conformed to an average value of 5 .times. 10.sup.7 per unit.
EXAMPLE 11
The processing system used to perform this example is set up in a manner
that generally corresponds to that shown above, with the difference in
this example pertaining to the red cell barrier filter assembly.
The red cell barrier filter assembly is configured in a manner that
generally corresponds to FIGS. 1 and 2. The housing, having a radially
positioned inlet and outlet, includes four ribs 8 on the inlet side, and,
on the outlet side, concentric channels 9a and eight radial channels 9b in
fluid communication with the outlet. The porous medium of the red cell
barrier filter assembly, positioned in the housing between the inlet and
the outlet, includes three zones of differing density, with the lowest
density at the upstream side of the medium, and increasing toward the
highest density at the downstream side of the medium. The first (upstream)
zone of the porous medium has a density of about 0.130 g/cc. The second
(middle) zone of the porous medium has a density of about 0.236 g/cc,
while the third (downstream) zone of the porous medium has a density of
about 0.294 g/cc.
The zones of the porous medium are preformed from 2.6 micron average
diameter PBT fibers, which have been surface modified in the manner as
described above and as taught in U.S. Pat. No. 4,880,548, using a mixture
of hydroxyethyl methacrylate and methacrylic acid in a monomer ratio of
0.35:1 to obtain a CWST of 95 dynes/cm and a zeta potential of -11.4
millivolts.
For each of the 20 tests summarized in this example, a human volunteer
donates a unit of whole blood, which passes through the needle line to be
collected in the collection bag (which already contains anticoagulant).
After mixing the blood with the anticoagulant in the collection bag, air
may be displaced into the needle line by stripping blood from the needle
line into the blood bag without releasing the stripper. The blood bag may
be oriented so that the remaining air bubble is just below the needle
line, and then the stripper may be released, and the needle line tubing
may be sealed, e.g., heat sealed.
Within approximately 8 hours after collection, the blood is processed as
described in the previous examples. As the PRP is expressed from the
collection bag, the red cell barrier assembly is held horizontally, with
the outlet of the assembly facing up, for priming. Once the PRP enters the
inlet of the assembly, the assembly may be laid down, if desired. PRP may
be expressed from the blood collection bag until red cells reach the
upstream surface of the porous medium, at which point the flow abruptly
stops, signalling the completion of filtration. The tubing from the outlet
side of the red cell barrier filter assembly may be clamped and heat
sealed, and the PRP bag may then be removed for further processing.
The PRP may be processed according to normal blood bank procedures to
create plasma and PC. Platelet counts may be taken and averaged for the 20
samples, and compared to the average platelet counts of 20 units of PC
prepared by conventional methods (i.e., without the red cell barrier
filter assembly) and obtained from a local blood bank. Using conventional
methods, the average platelet count may be about 6-7.times.10.sup.10
platelets per bag, while using the method according the instant invention
may yield a platelet count of about 9-9.5.times.10.sup.10 platelets per
bag, reflecting an increased yield of over 20%.
While the invention has been described in some detail by way of
illustration and example, it should be understood that the invention is
susceptible to various modifications and alternative forms, and is not
restricted to the specific embodiments set forth in the Examples. It
should also be understood that these Examples are not intended to limit
the invention but, on the contrary, the intention is to cover all
modifications, equivalents, and alternatives falling within the spirit and
scope of the invention.
Top